1. Field of the Invention
The present invention relates to a dual-state nutritional medium for the transport and maintenance of cells, and a method of using the same.
2. Description of Related Art
Growing and maintaining living cells outside the original host source is achieved by cell (or tissue) culture. Living cells outside the original host source may be maintained in vitro through numerous passages if prepared and provided with the appropriate nutrients. The nutrients required for the successful culture of cell lines have been established by many laboratories over at least the past fifty years. As a result, literally thousands of different cell lines have been initiated from the tissues of different species of animals, birds, insects, and plants, and are now in use in laboratories throughout the world.
Regardless of the cell system involved, the growth pattern of virtually all cells in culture is basically the same. Cell growth is dependent upon an appropriate nutrition and support system. Although cells may be grown successfully as a suspension, most cell cultures are preferentially grown as monolayers attached to a solid surface. This type of culture is particularly useful in the virus laboratory, but is also used in other disciplines. Cell cultures may be grown in either test tubes, flasks, or in wells of multi-well plastic plates. The most commonly used material for these containers is polystyrene. When appropriately treated, polystyrene supports the growth of cells in culture as monolayers on its surface.
Laboratories must utilize numerous cell lines from various origins for testing, since no single cell type is sensitive to all viruses. Many of the animal and plant cell cultures currently being used in laboratories are prepared in and obtained from other laboratories or commercial sources. The continued availability of cells to those laboratories without the capability of satisfying their cell culture needs is therefore very important. Such laboratories require cells from outside sources. These cells may be primary (first passage from the host) or serial (two or more passages from the original host). Regardless of passage number, to be useful, cell monolayers must be viable, albeit physiological changes from their in vivo state may occur.
At present, there are essentially two methods for obtaining monolayer cultures in multi-well plates for use in the laboratory. In the first method, cell cultures are transported as monolayers with a liquid medium filling the shipping container. In the second method, cell cultures are transported as cell suspensions. Both methods require that the shipping container be sealed to prevent loss of fluid. Neither method permits shipping of cells in open, multi-well plates or other unsealed containers, such as petri dishes.
Upon receipt of the cells by the end user, the cells must be seeded into appropriate culture plates and allowed to grow to the desired density before the cells can be used. This seeding and growth process requires an additional three to five days, requires additional effort on the part of the end user, and increases the possibility of cell contamination with extraneous microorganisms. The predominant use of cells is in the form of cultivated cell monolayers grown on the bottom of wells in polystyrene plates of various sizes (6, 12, 24, 48, or 96 wells per plate) previously seeded with cells in suspension. These plates come with a loosely fitting lid which allows gaseous exchange with the surrounding environment but which does not seal the plate. Once formed from the seeded cells, the cell monolayers must be covered with a liquid growth or maintenance medium. In this form, the monolayers in the wells are ready for use. Some cells may grow in forms other than monolayers, but if their growth is by adherence to the surface of the wells, the requirements for viable shipping needs are similar.
Currently, laboratories desiring such monolayers in multi-well plates must first obtain or produce cells in flasks or bottles. After disrupting the cell monolayer, the wells are seeded with cells and allowed to grow to a desired number (generally sufficient to cover the bottom of the well surface). Then, when ready (several days after seeding) the study material is inoculated. This procedure is both labor intensive and time consuming. If laboratories desiring such cell monolayers could obtain cell cultures in flat multi-well plates in a ready to use form, then much time and effort would be reduced. However, shipment of cell monolayers in wells using a liquid medium as a cover presents a problem, since shipment causes the liquid medium to spill and causes cellular disruption.
Attempts have been made to ship cell cultures in multi-well plates by sealing the wells with an adhesive or plastic sheet or plugging the wells with a stopper. These techniques have several drawbacks. A strip "sealing" cap is only practical for use in plates containing relatively small wells (e.g. those with 96 wells). Such a strip sealing cap is generally ineffective to prevent bubbles and other damage to cells caused by continuous fluid motion. The use of small plastic plugs as stoppers to stop the wells does not protect cells from damage due to bubbles and movement of the medium during transit. Plastic adhesive sheets are available for sealing the wells in a multi-well plate, but toxicity of the adhesive prevents their general use. These sheets also do not prevent bubble formation nor liquid movement during transport.
It is an object of the present invention to overcome the above-described difficulties and to provide a system and process by which cells may be transported and be ready for use by the end user virtually immediately upon receipt.